A plasma membrane preparation has been prepared from rat adipose tissue which has many of the features of the glucose transport system in intact cells. Plasma membranes isolated from cells exposed to insulin show an augmented D-glucose uptake as compared to membranes prepared from control adipocytes. Treatment of intact cells with anti-insulin serum (AIS) prior to cell rupture negated the stimulatory effect of insulin observed in the membranes, while treatment of subsequently isolated, insulin stimulated membranes with AIS did not lessen the observed stimulation of D-glucose transport. A number of spectroscopic techniques were used to assess the structure of the insulin-stimulated plasma membranes as compared to membranes from control adipocytes. By these techniques, no differences were observable between insulin stimulated and control membranes. It was of interest to see if the observed change in the plasma membrane glucose transport function might be due to the formation of a phosphoprotein. 32Pi was incubated with intact adipocytes to steady state 32P incorporation into protein (2 hours), and the cells were then ruptured and fractionated into the major subcellular fractions. The peptides in these fractions were separated on polyacrylamide gels containing sodium dodecylsulfate. 32P-phosphopeptides were assessed by quantitative radioautography. If the intact cells were exposed to epinephrine prior to rupture the phosphorylation of a large number of peptides, located in the cytosol, endoplasmic reticulum and plasma membrane was augmented. Insulin by itself stimulated the phosphorylation of a single species of MW 123,000 located in the cytosol and endoplasmic reticulum. Insulin added in the presence of epinephrine significantly inhibited certain epinephrine stimulated phosphorylation, but continued to stimulate the phosphorylation of the 123,000 dalton species. The effect of insulin on this latter species thus appears to be independent of cyclic AMP. Further studies are contemplated to try to better understand the processes by which hormone signals are transmitted to the enzymatic machinery of the cell. These will include an investigation of the changes in phospholipid and prostaglandin metabolism after hormone stimulation, as well as a search for an intracellular signal ("messenger") different from CAMP that may be an intermediate with the action of insulin.